Induction of Monocyte Chemoattractant Proteins in Macrophages via the Production of Granulocyte/Macrophage Colony-Stimulating Factor by Breast Cancer Cells.

Ji Ming Wang,Liangzhu Li,Tomozumi Imamichi,Teizo Yoshimura,Akihiro Matsukawa,Jonathan M. Weiss,Miwa Sato

doi:10.3389/fimmu.2016.00002

Abstract

Monocyte chemoattractant protein-1 (MCP-1)/CCL2 plays an important role in the initiation and progression of cancer. We previously reported that in 4T1 murine breast cancer, non-tumor stromal cells, including macrophages, were the major source of MCP-1. In the present study, we analyzed the potential mechanisms by which MCP-1 is upregulated in macrophages infiltrating 4T1 tumors. We found that cell-free culture supernatants of 4T1 cells (4T1-sup) markedly upregulated MCP-1 production by peritoneal inflammatory macrophages. 4T1-sup also upregulated other MCPs, such as MCP-3/CCL7 and MCP-5/CCL12, but modestly upregulated neutrophil chemotactic chemokines, such as KC/CXCL1 or MIP-2/CXCL2. Physicochemical analysis indicated that an approximately 2–3 kDa 4T1 cell product was responsible for the capacity of 4T1-sup to upregulate MCP-1 expression by macrophages. A neutralizing antibody against granulocyte/macrophage colony-stimulating factor (GM-CSF), but not macrophage CSF, almost completely abrogated MCP-1-inducing activity of 4T1-sup, and recombinant GM-CSF potently upregulated MCP-1 production by macrophages. The expression levels of GM-CSF in 4T1 tumors in vivo were higher than other tumors, such as Lewis lung carcinoma. Treatment of mice with anti-GM-CSF antibody significantly reduced the growth of 4T1 tumors at the injection sites but did not reduce MCP-1 production or lung metastasis in tumor-bearing mice. These results indicate that 4T1 cells have the capacity to directly upregulate MCP-1 production by macrophages by releasing GM-CSF; however, other mechanisms are also involved in increased MCP-1 levels in the 4T1 tumor microenvironment.

Highlights

Infiltration of leukocytes is observed in a number of human and mouse cancers [1, 2]
When peritoneal exudate cells (PEC) were co-cultured with 4T1 cells or cultured in the presence of 50% (v/v) 4T1-sup, the level of Monocyte chemoattractant protein-1 (MCP-1) in the culture supernatants was markedly increased in a manner dependent on the number of 4T1 cells (Figure 1A) or the concentration of 4T1-sup (Figure 1B). 4T1-sup did not increase the production of MCP-1 by TG-induced PEC from mice in which the MCP-1 gene was deleted in myeloid cells, confirming that myeloid cells, especially macrophages, were the source of MCP-1 in the culture (Figure 1C)
We previously reported that the chemokine MCP-1 produced by non-tumor cells in tumor stroma, both hematopoietic and non-hematopoietic cells, promoted spontaneous lung metastasis of 4T1 breast cancer cells [16]

Summary

Introduction

Infiltration of leukocytes is observed in a number of human and mouse cancers [1, 2]. The composition of tumorinfiltrating leukocytes and the role they play may vary in each tumor, they are generally immunosuppressive and provide a microenvironment that favors tumor growth. Monocyte chemoattractant protein-1 (MCP-1)/CCL2 is a chemokine with potent monocyte chemotactic activity. It was initially purified from the culture supernatant of a human malignant glioma [3] and a monocytic leukemic cell line [4], and is identical to the tumor-derived chemotactic factor, TDCF [5]. Accumulating evidence strongly suggests that the production of MCP-1 by tumors is responsible for the recruitment of immunosuppressive macrophages that promote tumor growth. MCP-1 is a candidate molecular target of cancer treatment [14]

Methods

Results

Conclusion